A recent surge of research revealed that there exist links between the gut microbiome and the central nervous system (microbiome-gut-brain axis). A recent surge of research has revealed that there are links between the gut microbiome and the central nervous system ("microbiome-gut-brain axis"). Changes in gut microbiota have been shown to alter mental well-being and potentially even have an impact on neurodegenerative diseases such Parkinson's disease [1, 2] and Alzheimer's disease [3]. Further connections have been demonstrated in germ-free mice, which exhibit altered brain synaptic [4]. Additionally, the administration of the probiotic L. rhamnosus (JB-1) to normal mice over the course of 28 days has resulted in changes in the chief inhibitory neurotransmitter, γ-Aminobutyric acid (GABA) receptor subtypes, which are accompanied by the presentation of anxiolytic behaviours [5]. These research efforts highlight the important role of the gut microbiome in the bi-directional communication of the gut-brain axis which appears to be mediated by the vagus nerve. Despite the lack of understanding of the underlying mechanism of interaction there exists the potential for using probiotics as therapeutic adjuncts in major depressive disorders. This study focuses on the development and application of magnetic resonance spectroscopy (MRS) for the tracking of neuro-chemical changes associated with probiotic bacteria exposure in animal models.Experiments were carried out in twenty 28 adult male BALB/c mice (Charles River, Wilmington MA) weighing 25–35 g. Mice were gavaged with 200mL of phosphate buffered saline (PBS) - control group, n=14, or with 200mL of re-suspended L. rhamnosus (JB-1), 1 × 109 cfu (JB-1, group n=14) daily. Treatment with JB-1 lasted 4 weeks. MRS (single voxel) was performed for all animals before the start of gavaging (week 0), during JB-1 exposure (weeks 1, 2, 3, 4 for n=11, 6 control and 5 JB-1 treated and weeks 2, 4 and 8 for n=17, 8 control and 9 JB-1 treated). Proton MRS spectra were acquired for each animal over the region of interest using point resolved spectroscopy (PRESS). The following sequence parameters were: (repetition time/echo time=2500/17.5ms, number of transients=512, bandwidth=3000Hz, data points= 4096). The water signal was suppressed using eight variable power RF pulses with optimized relaxation delays (VAPOUR). VAPOUR inter-pulse delays and pulse amplitudes were optimized manually for each animal to achieve optimal water suppression. Additional non water suppressed spectra were acquired to allow for the normalization of neuro-metabolite concentrations to the concentration of in vivo brain water.ROI and representative spectra are shown in Fig. 1. Statistically significant (p<0.01) changes in concentrations of GABA, glutamate/glutamine and NAA were observed four weeks after the start of treatment in the hippocampus of mice treated with the probiotic (Fig.2). Anatomical scan could only be collected in sixteen animals and showed no change in the volume of the hippocampus with exposure to probiotic (Fig. 3). Glutamate and NAA levels remained elevated 2 weeks after the end of treatment despite the bacteria being non-colonizing. GABA levels returned to baseline levels. Glutamate concentration levels measure by ex-vivo ELISA correlated with in-vivo measurements (P<0.05, R=0.77). This initial experiment demonstrates that there appears to be an effect on neuro-transmitter levels resulting from probiotic exposure and that the effect size is large enough to be observed using MRS.